Rickettsia rickettsii is the causative agent of Rocky Mountain spotted fever. Strains of R. rickettsii and other species of rickettsiae differ considerably in virulence. We are applying modern molecular biological techniques to define and characterize these strains and attempting to identify virulence determinants on these obligate intracellular parasites. Current efforts are focused on a pair of high molecular weight surface proteins. These immunodominant surface protein antigens have estimated mol. masses of 190 and 135 kDa and are term rOmp A and B, respectively. The DNA sequence of the gene encoding the rOmp B protein of R. rickettsii has been completed to identify a gene with the capacity to specify a protein of approximately 168 kDa. N-terminal amino acid sequencing of a 32 kDa outer membrane protein revealed that it is encoded by the 3' terminal end of he rOmp B gene suggesting that the rOmp B protein is proteolytically processed to yield the mature 135 kDa rOmp B protein and a 32 kDa fragment. Both polypeptides seem to be stable products and remain non-covalently associated on the outer surface of rickettsiae. Although we have not yet detected the rOmp B precursor on virulent R. rickettsii, analysis of an avirulent mutant of R. rickettsii revealed an absence of the 32 kDa fragment with a correspondingly larger rOmp B protein as would be predicted from an inability to cleave the expected precursor. The 190 kDa rOmp A protein of R. rickettsii (R strain) contains a large region of tandemly arranged repeats. Using a clone encoding this repeat region as a probe, homologous sequences were discovered within the genomes of all but one species of spotted fever group rickettsiae tested. Southern blot analysis on DNA fragments internal to a 3.8 kb Pstl segment showed polymorphisms between species of R. rickettsii. Analysis of the promoter regions of the rOmp A and B genes by cloning the promoter in front of a chloramphenicol- acetyl transferase reporter gene revealed that these rickettsial promoters are active in E. coli. the activity of the promoters approximates the relative abundance of the respective proteins on rickettsia. This information will provide a rational basis for eventual attempts to express rickettsial genes in alternate hosts. Expression of rOmp A and B in recombinant hosts will be a powerful tool in definition of function.